Zheng L M, Wang L N, Liang C, Peng C J, Tang W Y, Zhang X L, Li Y, Tang Y L, Huang L B, Luo X Q
Pediatric Department, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China.
Zhonghua Xue Ye Xue Za Zhi. 2020 Oct 14;41(10):836-842. doi: 10.3760/cma.j.issn.0253-2727.2020.10.008.
Endoplasmic reticulum stress(ERS)was used as the research emphasis to further investigate the mechanisms of apoptosis of FLT3-ITD-mutated leukemia cells and decreased expression of FLT3-ITD mutated protein induced by all-trans retinoic acid(ATRA). FLT3-ITD-mutated leukemia cell lines(MV4-11 and MOLM13)were treated with ATRA. Flow cytometry was conducted to assess cell apoptosis. Real-time fluorescent quantitative PCR(RT-qPCR)and Western blot were used to detect the expression of ERS-related and autophagy-related genes and protein, respectively. A low-dose ATRA further increased FLT3-ITD cells and ERS levels. ATRA acted on the ERS-related PERK/eif2ɑ signaling pathway and continued to increase the ERS of FLT3-ITD cells, resulting in an upregulation of apoptotic gene CHOP expression. After the treatment with ATRA, FLT3-ITD protein in FLT3-ITD cells was decreased. Of the two main ERS-related protein degradation pathways, ER-associated degradation(ERAD)and ER-activated autophagy(ERAA), the expression of ERAD-related protein ATF6 in FLT3-ITD cells was not significantly changed on ATRA, whereas the expression of ERAA-related proteins Atg7 and Atg5 were significantly increased. ATRA further raises the ERS level of FLT3-ITD cells continuously by activating the ERS-related PERK/eif2ɑ signal pathway and induces FLT3-ITD protein autophagy degradation through ERAA pathway, which induces apoptosis of FLT3-ITD-mutated leukemia cells. These results provide preliminary evidence on the use of ATRA in the treatment of refractory leukemia with FLT3-ITD.
以内质网应激（ERS）为研究重点，进一步探讨全反式维甲酸（ATRA）诱导FLT3-ITD突变白血病细胞凋亡及FLT3-ITD突变蛋白表达降低的机制。用ATRA处理FLT3-ITD突变白血病细胞系（MV4-11和MOLM13）。采用流式细胞术评估细胞凋亡。分别用实时荧光定量PCR（RT-qPCR）和蛋白质免疫印迹法检测ERS相关基因和自噬相关基因及蛋白的表达。低剂量ATRA进一步增加了FLT3-ITD细胞及ERS水平。ATRA作用于ERS相关的PERK/eif2ɑ信号通路，持续增加FLT3-ITD细胞的ERS，导致凋亡基因CHOP表达上调。经ATRA处理后，FLT3-ITD细胞中的FLT3-ITD蛋白减少。在ERS相关的两条主要蛋白降解途径，即内质网相关降解（ERAD）和内质网激活自噬（ERAA）中，ATRA处理后FLT3-ITD细胞中ERAD相关蛋白ATF6的表达无明显变化，而ERAA相关蛋白Atg7和Atg5的表达显著增加。ATRA通过激活ERS相关的PERK/eif2ɑ信号通路持续进一步提高FLT3-ITD细胞的ERS水平，并通过ERAA途径诱导FLT3-ITD蛋白自噬降解，从而诱导FLT3-ITD突变白血病细胞凋亡。这些结果为ATRA用于治疗FLT3-ITD难治性白血病提供了初步证据。
